Our previous work demonstrated that functional voltage-dependent and ligand-activated channels mature during early differentiation of amphibian spinal neurons, with a similar time course both in vivo and dissociated cell culture. These findings suggest that ion channel activity participates in signal transduction that influences subsequent steps of development. Our recent work indicates that spontaneous transient elevations of intracellular calcium affect later aspects of differentiation in vitro, in both neurons and myocytes. The proposed studies address the role and mechanisms by which ion channel activity controls long-lasting molecular events required for proper differentiation, both in vitro and in vivo. The proposed research has three specific aims. The first two address different aspect of calcium-dependent regulation of neurite extension and navigation. The third aim analyzes calcium-dependent expression of neurotransmitter. Our first aim is to determine the function of spontaneous calcium transients in neurite extension and navigation in vivo. Our second aim is to analyze the roles of glutamate receptors in elevating intracellular calcium and regulating neurite outgrowth in vivo. Our third aim is to analyze the basis of calcium-dependent regulation of expression of GABA by spontaneous transient neuronal calcium spikes in culture. The immediate goal is to test hypotheses about specific mechanisms underlying differentiation of vertebrate spinal neurons in order to define the roles of early ion channel activity in driving differentiation. The long term goal is to provide information about the cellular and molecular machinery that governs processes of development. It is expected that this work will contribute to understanding developmental disorders of the nervous system.